This year emphasis has been shifted towards molecular biological studies with several major lines of investigation being pursued to identify, 1) autoantibodies in insulin-dependent diabetes mellitus (IDDM), 2) activated oncogenes in human insulinomas and other endocrine diseases, and 3) genes essential for beta-cell function. Recombinant DNA expression libraries of cDNA from a rat insulinoma cell line have been screened with autoimmune serum from IDDM patients and several clones coding for potential autoantigens have been isolated. Our attempts to establish human beta-cell lines by transfection and microinjection with oncogenes have met with preliminary, success. These cell lines will be invaluable for assessing the immunological status of IDDM patients. We are well into the characterization of one possibly two activated oncogenes from human insulinomas. DNA sequences responsible for tumorigenesis have been isolated after several rounds of tumor formation in nude mice and seem to be unrelated to any known oncogene. Nucleotide sequence analyses and tumorigenesis will be used to determine whether we have isolated a new oncogene, associated with tumors of beta-cell origin. We have initiated a second project to isolate transforming genes from other endocrine diseases, such as thyroid carcinoma and multiple endocrine neoplasia. In a search for genes essential in beta-cell function we have focused on the superoxide dismutase (SOD) gene, responsible for oxygen detoxification. This gene is involved in several forms of chemically-induced diabetes and is overexpressed in beta-cells. We have cloned, isolated and sequenced cDNA copies of rat SOD mRNA, and we are presently studying its transcriptional status in insulin-producing cells. Finally, we are developing a gene therapy model of intervention in hormone deficiency diseases. We have constructed recombinant plasmids that carry the rat insulin I gene under the regulation of the MMTV LTR and are studying glucocorticoid-modulation of insulin expression in lymphocytes transfected with this construct.